Low-tyramine stevia plant

Abstract

The present invention provides a low tyramine-content Stevia plant having a lower tyramine content as compared with the wild type Stevia species. The present invention also provides a method of producing such a low tyramine-content Stevia plant, and an extract and a steviol glycoside purified product obtainable from such a plant.

1 . A low tyramine-content  Stevia  plant having a tyramine content of less than 0.092% per unit mass of a dried leaf. 
     
     
         2 . The plant according to  claim 1 , having a rebaudioside D content of 1% or more per unit mass of a dried leaf. 
     
     
         3 . The plant according to  claim 1  or  2 , wherein the plant is homozygous for the allele wherein the base at the position corresponding to position 201 of SEQ ID NO: 150 is C. 
     
     
         4 . The plant according to any one of  claims 1  to  3 , wherein the plant is homozygous for the allele wherein the base at the position corresponding to position 201 of SEQ ID NO: 1 is A. 
     
     
         5 . The plant according to any one of  claims 1  to  4 , further having at least one of the following genetic features.
 (1) Homozygous for the allele wherein the base at the position corresponding to position 40 of SEQ ID NO: 2 is T. 
 (2) Homozygous for the allele wherein the base at the position corresponding to position 44 of SEQ ID NO: 3 is T. 
 (3) Homozygous for the allele wherein the base at the position corresponding to position 41 of SEQ ID NO: 4 is C. 
 (4) Homozygous for the allele wherein the portion corresponding to positions 55-72 of SEQ ID NO: 5 is deleted. 
 
     
     
         6 . The plant according to any one of  claims 1  to  5 , wherein the plant is heterozygous for the allele wherein the base at the position corresponding to position 49 of SEQ ID NO: 6 is A. 
     
     
         7 . The plant according to any one of  claims 1  to  6 , wherein the plant is a non-genetically modified plant. 
     
     
         8 . The plant according to any one of  claims 1  to  7 , wherein the plant includes a  Stevia  plant subjected to a mutagenesis treatment and a progeny plant thereof. 
     
     
         9 . A seed, a tissue, a tissue culture or a cell of the plant according to any one of  claims 1  to  8 . 
     
     
         10 . The tissue, tissue culture or cell according to  claim 9 , which is selected from an embryo, a meristem cell, a pollen, a leaf, a root, a root apex, a petal, a protoplast, a leaf section and a callus. 
     
     
         11 . A method of producing a low tyramine-content  Stevia  plant having a tyramine content of less than 0.092% per unit mass of a dried leaf, the method comprising a step of crossing the plant according to any one of  claims 1  to  8  with a second  Stevia  plant. 
     
     
         12 . The method according to  claim 11 , wherein the second plant is the plant according to any one of  claims 1  to  8 . 
     
     
         13 . An extract of the plant according to any one of  claims 1  to  8 , or of the seed, tissue, tissue culture or cell according to  claim 9  or  10 , wherein the extract has a low tyramine content. 
     
     
         14 . A method of producing a low tyramine-content  Stevia  extract, comprising a step of obtaining an extract from the plant according to any one of  claims 1  to  8 , or from the seed, tissue, tissue culture or cell according to  claim 9  or  10 . 
     
     
         15 . A method of producing a low tyramine-content steviol glycoside purified product, comprising a step of purifying steviol glycosides from the  Stevia  extract according to  claim 13 . 
     
     
         16 . The method according to  claim 15 , wherein the steviol glycosides include rebaudioside A, rebaudioside B, rebaudioside C, rebaudioside D, rebaudioside E, rebaudioside F, rebaudioside M, rebaudioside N, rebaudioside O, stevioside, steviolbioside, rubusoside, dulcoside A or a combination thereof. 
     
     
         17 . A low tyramine-content steviol glycoside purified product produced by the method according to  claim 15  or  16 . 
     
     
         18 . A low tyramine-content food or beverage, sweetener composition, flavor or medicament comprising the extract of the tissue, tissue culture or cell according to  claim 13 , or the purified product according to  claim 17 . 
     
     
         19 . A method of producing a food or beverage, a sweetener composition, a flavor or a medicament, comprising:
 a step of providing the extract of the tissue, tissue culture or cell according to  claim 13 , or the purified product according to  claim 17 ; and   a step of adding the extract or the purified product to a raw material for the food or beverage, sweetener composition, flavor or medicament.   
     
     
         20 . A method of screening for a low tyramine-content  Stevia  plant, comprising a step of detecting from the genome of a test  Stevia  plant the presence and/or the absence of a genetic feature of being homozygous for the allele wherein the base at the position corresponding to position 201 of SEQ ID NO: 150 is C. 
     
     
         21 . The method according to  claim 20 , further comprising a step of detecting from the genome of a test  Stevia  plant the presence and/or the absence of a genetic feature of being homozygous for the allele wherein the base at the position corresponding to position 201 of SEQ ID NO: 1 is A. 
     
     
         22 . The method according to  claim 20  or  21 , further comprising a step of detecting from the genome of a test  Stevia  plant the presence and/or the absence of the following genetic features (1) to (4).
 (1) Homozygous for the allele wherein the base at the position corresponding to position 40 of SEQ ID NO: 2 is T. 
 (2) Homozygous for the allele wherein the base at the position corresponding to position 44 of SEQ ID NO: 3 is T. 
 (3) Homozygous for the allele wherein the base at the position corresponding to position 41 of SEQ ID NO: 4 is C. 
 (4) Homozygous for the allele wherein the portion corresponding to positions 55-72 of SEQ ID NO: 5 is deleted. 
 
     
     
         23 . The method according to any one of  claims 20  to  22 , further comprising a step of detecting from the genome of a test  Stevia  plant the presence and/or the absence of a genetic feature of being heterozygous for the allele wherein the base at the position corresponding to position 49 of SEQ ID NO: 6 is A. 
     
     
         24 . The method according to any one of  claims 20  to  23 , wherein the step of detecting a genetic feature is performed by use of CAPS method, dCAPS method or TaqMan PCR method. 
     
     
         25 . The method according to any one of  claims 20  to  24 , further comprising a step of measuring the content of a tyramine in a test  Stevia  plant tissue. 
     
     
         26 . A screening kit for a low tyramine-content  Stevia  plant, comprising a reagent for detecting the presence and/or the absence of a genetic feature of being homozygous for the allele wherein the base at the position corresponding to position 201 of SEQ ID NO: 150 is C. 
     
     
         27 . The kit according to  claim 26 , further comprising a reagent for detecting the presence and/or the absence of a genetic feature of being homozygous for the allele wherein the base at the position corresponding to position 201 of SEQ ID NO: 1 is A. 
     
     
         28 . The kit according to  claim 26  or  27 , further comprising a reagent for detecting the presence and/or the absence of the following genetic features (1) to (4).
 (1) Homozygous for the allele wherein the base at the position corresponding to position 40 of SEQ ID NO: 2 is T. 
 (2) Homozygous for the allele wherein the base at the position corresponding to position 44 of SEQ ID NO: 3 is T. 
 (3) Homozygous for the allele wherein the base at the position corresponding to position 41 of SEQ ID NO: 4 is C. 
 (4) Homozygous for the allele wherein the portion corresponding to positions 55-72 of SEQ ID NO: 5 is deleted. 
 
     
     
         29 . The kit according to any one of  claims 26  to  28 , further comprising a reagent for detecting the presence and/or the absence of a genetic feature of being heterozygous for the allele wherein the base at the position corresponding to position 49 of SEQ ID NO: 6 is A. 
     
     
         30 . The kit according to any one of  claims 26  to  29 , wherein the reagent comprises a primer and/or a probe for use in CAPS method, dCAPS method or TaqMan PCR method. 
     
     
         31 . A method of producing a low tyramine-content  Stevia  plant, comprising a step of introducing a variation from T to C to a position corresponding to position 201 of SEQ ID NO: 150. 
     
     
         32 . The method according to  claim 31 , wherein the introduction of the variation is performed by a mutagenesis treatment.